The Ras proteins (Ha-Ras, Ki4a-Ras, Ki4b-Ras and N-Ras) are part of a signaling pathway that links cell surface growth factor receptors to nuclear signals initiating cellular proliferation. Biological and biochemical studies of Ras action indicate that Ras functions like a G-regulatory protein. In the inactive state, Ras is bound to guanosine diphosphate (GDP). Upon growth factor receptor activation Ras is induced to exchange GDP for guanosine triphosphate (GTP) and undergoes a conformational change. The GTP-bound form of Ras propagates the growth stimulatory signal. The signal is terminated by the intrinsic GTPase activity of Ras, facilitated by the GTPase activating protein (GAP), which returns the protein to its inactive GDP bound form (D. R. Lowy and D. M. Willumsen, Ann. Rev. Biochem. 62:851–891 (1993)). Mutated ras genes (Ha-ras, Ki4a-ras, Ki4b-ras and N-ras) are found in many human cancers, including colorectal carcinoma, exocrine pancreatic carcinoma, and myeloid leukemias. The protein products of these genes are defective in their GAP-assisted GTPase activity and transmit an uncontrolled growth stimulatory signal.
Ras must be localized to the plasma membrane for both normal and oncogenic functions. At least 3 post-translational modifications are involved with Ras membrane localization, and all 3 modifications occur at the C-terminus of Ras. The Ras C-terminus contains a sequence motif termed a “CAAX” or “Cys-Aaa.sup.1-Aaa.sup.2-Xaa” box (Cys is cysteine, Aaa is an aliphatic amino acid, the Xaa is any amino acid) (Willumsen et al., Nature 310:583–586 (1984)). Depending on the specific sequence, this motif serves as a signal sequence for the enzymes farnesyl-protein transferase or geranylgeranyl-protein transferase, which catalyze the alkylation of the cysteine residue of the CAAX motif with a C15 or C20 isoprenoid, respectively. (S. Clarke., Ann. Rev. Biochem. 61:355–386 (1992); W. R. Schafer and J. Rine, Ann. Rev. Genetics 30:209–237 (1992)). The Ras protein is one of several proteins that are known to undergo post-translational farnesylation. Other farnesylated proteins include the Ras-related GTP-binding proteins such as Rho, fungal mating factors, the nuclear lamins, and the gamma subunit of transducin. James, et al., (J. Biol. Chem. 269, 14182 (1994)) have identified a peroxisome associated protein, Pxf, which is also farnesylated. James, et al. have also suggested that there are farnesylated proteins of unknown structure and function in addition to those listed above.
Inhibition of farnesyl-protein transferase (FPTase) has been shown to block the growth of Ras-transformed cells in soft agar and to modify other aspects of their transformed phenotype. It has also been demonstrated that certain inhibitors of FPTase selectively block the processing of the Ras oncoprotein intracellularly (N. E. Kohl et al., Science, 260:1934–1937 (1993) and G. L. James et al., Science, 260:1937–1942 (1993). Recently, it has been shown that an inhibitor of FPTase blocks the growth of ras-dependent tumors in nude mice (N. E. Kohl et al., Proc. Natl. Acad. Sci U.S.A., 91:9141–9145 (1994)) and induces regression of mammary and salivary carcinomas in ras transgenic mice (N. E. Kohl et al., Nature Medicine, 1:792–797 (1995)).
It has also been reported that FPTase inhibitors are also inhibitors of proliferation of vascular smooth muscle cells and are, therefore, useful in the prevention and therapy of arteriosclerosis and diabetic disturbance of blood vessels (Japanese Patent H7-112930).
Inhibitors of FPTase have been classified in two general classes. The first are analogs of farnesyl diphosphate (FPP), while the second class of inhibitors is structurally related to the enzyme's protein substrates (e.g., Ras). The peptide derived inhibitors that have been described are generally cysteine-containing molecules that are related to the CAAX motif that is the signal for protein prenylation. (Schaber et al., ibid; Reiss et. al., ibid; Reiss et al., PNAS, 88:732–736 (1991)). Such inhibitors may inhibit protein prenylation while serving as alternate substrates for the FPTase enzyme, or may be purely competitive inhibitors (U.S. Pat. No. 5,141,851, University of Texas; N. E. Kohl et al., Science, 260:1934–1937 (1993); Graham, et al., J. Med. Chem., 37, 725 (1994)). In general, deletion of the thiol from a CAAX derivative has been shown to dramatically reduce the inhibitory potency of the compound. However, the thiol group potentially places limitations on the therapeutic application of FPTase inhibitors with respect to pharmacokinetics, pharmacodynamics and toxicity. Therefore, a functional replacement for the thiol is desirable.
It has recently been disclosed that certain tricyclic compounds which optionally incorporate a piperidine moiety are inhibitors of FPTase (WO 95/10514, WO 95/10515 and WO 95/10516). Imidazole-containing inhibitors of FPTase have also been disclosed (WO 95/09001 and EP 0 675 112 A1). A number of benzodiazepine-based FPTase inhibitors are also described in U.S. Pat. No. 6,011,029, which is commonly assigned with this application.
There is, therefore, a need for a variety of pharmaceutically useful FPTase inhibitors, as well as for safer and more efficient processes for their manufacture.